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D-Serine in animal models of substance abuse, anxiety and stroke - Joseph T Coyle

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Watching at the "D" side: D-amino acids and their significance in neurobiology
June 05 -June 09, 2016 – Lake Como School of Advanced Studies

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D-Serine in animal models of substance abuse, anxiety and stroke - Joseph T Coyle

  1. 1. D-Serine in animal models of substance abuse, anxiety and stroke Joseph T Coyle, MD Eben S draper Professor of Psychiatry Harvard Medical & McLean Hospital Lake Como School of Advanced Studies Watching at the “D” side: D-amino acids and ther significance in neurobiology
  2. 2. 6/9/16 2 Where are serine racemase and D-serine localized in brain?
  3. 3. 6/9/16 3 The Confusing Immunocytochemical Localizations of Serine Racemase and D-Serine • D-serine is localized to astrocytes (“glial transmitter”): Schell MJ et al., J Neurosci. 92: 3048-52, 1995. • SR and D-serine are also localized to cultured neurons: Kartvelishvily E, et al., J.Biol Chem. 2006 May 19;281(20):14151-62. • SR and D-serine are localized only to forebrain glutamatergic and GABAergic neurons: Miya K, et al., J Comp Neurol. 2008 Oct 20;510(6):641-54.
  4. 4. 6/9/16 4 Miya et al. demonstrate serine racemase in neurons Miya et al., J comp Neurol 510: 641-510, 2008
  5. 5. Miya et al., demonstrate that serine racemase is not expressed in astrocytes in adult mouse brain Miya et al., J comp Neurol 510: 641-510, 2008
  6. 6. 6/9/16 66/9/16 66/9/16 6 Serine Racemase is Expressed Primarily in Pyramidal Glutamatergic Neurons and GABAergic Inter-neurons in Forebrain: Results of Cell-Specfic Gene Inactivation PCR Genotyping: 1. SR+/+ 2. SRfl/+ 3. SRfl/fl 4. Cre transgene 5. No Cre transgene Benneyworth et al., 2012; Takagi et al., in preparation
  7. 7. 0.5 mM L-serine 10 mM L-serine High blocking L-serine prevents artifactual immuno-staining for D-serine with anti-D- serine antiserum Balu DT, Takagi S, Puhl MD, Benneyworth MA and Coyle, JT. Cell Mol Neurobiol 34: 419-435, 2014.
  8. 8. 6/9/16 8 D-Serine Does Not Co-Localize with Astrocyte Markers, GFAP and S100β Balu et al., Cell Mol Neurobiol 34: 419-35, 2014
  9. 9. 6/9/16 9 D-Serine Co-Localizes with Cortical GABAergic Neurons Balu et al., Cell Mol Neurobiol 34: 419-35, 2014
  10. 10. 6/9/16 10 Neuronal but not astrocyte silencing of serine racemase expression attenuates long-term potentiation at hippocampal CA1 synapse Benneyworth et al., Cell Mol Neurobiol 32: 613-24, 2012 Ishikawa et al., J Neural Transm 122: 1099-103, 2015
  11. 11. 6/9/16 11 Serine Racemase is expressed in some forms of astrocytes !
  12. 12. 12 Serine Racemase is expressed in cultured astrocytes (Type II) and synthesizes D-serine S Li, H Wolosker, JT Coyle, in preparation Abe et al., J Cereb Blood Flow& Metab 34: 1928-35, 2014
  13. 13. 6/9/16 136/9/16 136/9/16 13 Serine Racemase is Expressed in Reactive Astrocytes After Traumatic Brain Injury D Balu, E Perez-Martinez2, S Takagi, JT Coyle and DJ Liebl in preparation
  14. 14. 6/9/16 14 We find reactive gliosis consists of a rapid, but quickly attenuated, induction of gene expression after insult and identify induced Lcn2 and Serpina3n as strong markers of reactive astrocytes. Strikingly, reactive astrocyte phenotype strongly depended on the type of inducing injury…. Reactive astrocytes in ischemia exhibited a molecular phenotype that suggests that they may be beneficial or protective, whereas reactive astrocytes induced by LPS exhibited a phenotype that suggests that they may be detrimental. Zamanian et al., J Neurosci 32: 6391-6410, 2012 Ben Bares shows that the type of insult affects gene expression in reactive astrocytes
  15. 15. Source of artifacts with regard to the localization of D-serine and serine racemase 1. High concentrations of L-serine in astrocytes result in immuno-cross- reactivity with D-serine-BSA antiserum. Solution: increase L-serine- BSA blocking concentration and use SR-/- to determine immuno- specificity. 2. Immuno-crossreactivity of serine racemase antiserum: Solution: use SR-/- tissue to ensure immuno-specificity. (Santa Cruz: “antibody suitable for immunocytochemistry but not western blots due multiple bands” !!??). Use SR-/- as controals 3. The astrocyte toxin fluroacetate decreases synaptic D-serine. Problem: Astrocytes are the source of L-serine for D-serine synthesis in astrocytes (Serine shuttle). 4. Cultured astrocytes as a model for studying serine racemase regulation. Solution: cultured astrocytes do not represent Type I but rather a form of Type II astrocytes
  16. 16. 6/9/16 16 Don’t forget glycine! Evoked NMDA currents PrePulse Inhibition MWM probe task spent 4X more time over missing platform than wild type (p<0.001) Tsai et al., PNAS 101: 8489, 2004
  17. 17. 6/9/16 17 Glutamtergic Gabaergic A modest proposal: D-serine is an auto-modulator to ensure immediate activation of NMDA receptors with glutamate release; Glycine is recruited on hig demand. Benneywort et al.,Cell Molec Neurobiol 32: 32-61, 2012 Li et al., Nature Comm. DOI: 10.1038/ncomms2779 Lin et al., Fron Cell Neurosci. 10:34. doi: 10.3389/fncel.2016.00034.
  18. 18. 6/9/16 18 Addiction and D-serine
  19. 19. 6/9/16 19 Neuroplastic changes triggered by drugs have been uncovered not only in the nucleus accumbens (a crucial brain-reward region) but also in the dorsal striatum (a region implicated in the encoding of habits and routines), the amygdala (a region involved in emotions, stress, and desires), the hippocampus (a region involved in memory), and the prefrontal cortex (a region involved in self-regulation and the attribution of salience [the assignment of relative value]). All these regions of the brain participate in the various stages of addiction, including conditioning and craving. Volkow ND, Koob GF and McLellan AT, N Engl J Med 374: 363-71, 2016 Substance use disorders involve neuroplastic alterations in several brain regions mediating reward, memory and emotions
  20. 20. 6/9/16 20 Neuroplasticity drives addiction J Peters & TJ DeVries: Cold Spring Harb 2: 1-13, 2012
  21. 21. 6/9/16 21 • Substance abuse disorders are 3-5 times more prevalent in individuals with schizophrenia (Coyle, 2006; Kavanaugh et al., 2002; Regier et al., 1990) • Comprises all classes of abused substances (nicotine, stimulants, alcohol and opiates)and is associated with poor schizophrenia treatment outcomes (Cantor-Graae et al., 2001; Erkiran et al., 2007; Green, 2007) Co-morbid schizophrenia and substance abuse disorders
  22. 22. 6/9/16 22 Acquisition and expression of behavioral sensitization to amphetamine in WT and SR-/- mice Benneyworth and Coyle, Neuropsychopharmacology 37: 2496-2504, 2012
  23. 23. 6/9/16 23 Benneyworth and Coyle, Neuropsychopharmacology 37: 2496-2504, 2012 Impaired extinction of amphetamine sensitization is associated with lack of reduction in spine density in the core of the nucleus accumbens
  24. 24. 6/9/16 24 Day  1:  Habituation Saline  injection;;  free  access  (Plexi floor) Days  2-­3:  Conditioning AM: Saline  injection;;  confined  to  CS-­ side PM: Cocaine  injection;;  confined  to  CS+   side Day  4:  Test Saline  injection;;  free  access Days  5-­16:  Extinction Saline  injection;;  free  access Day  17:  Reinstatement Cocaine  injection;;  free  access Effects of D-serine and glycine availability on conditioned place preference (CPP) to cocaine, a model of craving Puhl et al., J Pharma Exp Ther 353: 465-470, 2015
  25. 25. 6/9/16 25 Conditioned Place Preference Chambers
  26. 26. 6/9/16 26 Effect of SR-/- and GlyT1+/- on acquisition and extinction of cocaine –induced conditione place preference Puhl et al., J Pharma Exp Ther 353: 465-470, 2015
  27. 27. 6/9/16 27 Puhl et al., J Pharma Exp Ther 353: 465-470, 2015 Effect of pharmacologic reversal of SR-/- and GlyT1+/- on cocaine-induced place preference Gavestinal=NMDAR glycine modulatory site antagonist D-serine treatment normalizes hippocampal levels of D-serine
  28. 28. 6/9/16 28 Reduced locomoter sensitization to cocaine in SR-/- mice Puhl et al., J Pharma Exp Ther 353: 465-470, 2015
  29. 29. 6/9/16 29 Electrical  stimulation  of  excitatory   inputs  of  the  mesocorticolimbic dopamine  system (Bielajew &   Shizgal,  1986;;  Yeomans et  al.,  2000) Targets  medial  forebrain  bundle   (MFB) at  the  level  of  the  lateral   hypothalamus Intracranial self-stimulation Nieuwenhuys et  al.,  J  comp  Neurol 206:  49-­81,  1982 *
  30. 30. 6/9/16 30 Advantages MFB  stimulation  is  extremely  rewarding at  relatively   low  current  intensities  (Carlezon &  Chartoff,  2007;;   Liebman,  1983;;  Roybal et  al.,  2007) Little  or  no motor  side  effects  (Liebman,  1983) Highly  sensitive  assay  of  hedonic  state (Markou &   Koob,  1991;;  Tomasiewicz et  al.,  2006) Drugs  of  abuse  enhance ICSS  responding,  while   treatments  that  produce  dysphoria (e.g.,  withdrawal,   stress,  etc.)  attenuate ICSS  responding  (Muschamp et   al.,  2012) Intra-cranial self-stimulation
  31. 31. 6/9/16 31 Protocol Rate-frequency (RF) 1-h to 1.5-h sessions Fixed current (from TR) 15 frequencies in descending order (0.05 log10 unit steps), 1 minute per frequency Find frequency (T0) at which stimulation becomes rewarding (i.e., “threshold”) ~4-6 weeks (weekends off) Intra-cranial self-stimulation Carlezon WA Jr, Chartoff EHIntracranial self-stimulation (ICSS) in rodents to study the neurobiology of motivation. Nat Protoc. 2007;2(11):2987-95.
  32. 32. 6/9/16 32 Cocaine does not attenuate ICSS in SR-/- mice like in WT Puhl et al., in preparation
  33. 33. 6/9/16 33 SR-/- mice exhibit increased dopamine release in the nucleus accumbens core at base-line and with cocaine Puhl et al., in preparation SR-/- male mice exhibit hyperactivity and reareing, consistent with increased dopamine release
  34. 34. 6/9/16 34 D-Serine is the NMDA receptor co-agonist in the nucleus accumbens core Curcio et al., Brain 136: 1216-30, 2013
  35. 35. 6/9/16 35 Chronic cocaine injections reduces SR expression, D-serine release and increased D-amino acid oxidase expression in the nucleus accumbens core Curcio et al., Brain 136: 1216-30, 2013
  36. 36. 6/9/16 36 Infusion of D-serine into the rat nucleus accumbens increases extinction Liu et al., Pharmacology, Biochemistry and Behavior 143: 44-56, 2016
  37. 37. 6/9/16 37 Infusion of D-serine into the nucleus accumbens blocks locomotor sensitization in the rat D’Ascenzo et al., Fontiers in Synaptic Neuroscience 6: 1-6, 2014
  38. 38. 6/9/16 38 Infusion of D-serine into the nucleus accumbens enhances the aversive effects of quinine in compulsive alcohol alcohol consuming rats mediated by medial prefrontal cortex afferents Self et al., Nat Neurosci 16:1094-1100, 2013 Self et al., Neuropsychopharmacology (2015) advance online publication
  39. 39. 6/9/16 39 Anxiety Disorders
  40. 40. 6/9/16 40 Neuronal circuits of fear expression Dejean et al., Biol Psychtr 78: 298-306, 2015
  41. 41. 6/9/16 41 Rauch et al., Biol Psychiatry 47: 769-776 Subjects with post-traumatic stress disorder show an exaggerated amygdala responses
  42. 42. 6/9/16 42 Trace Fear Conditioning Pine DS, Fox NA. Annu Rev Psychol. 2015 Jan 3;66:459-85.
  43. 43. 6/9/16 436/9/16 436/9/16 43 SR-/- mice exhibit deficits in trace fear conditioning that are reversed by restoring hippocampal D-serine levels in adulthood Balu et al., Proc Natl Acad Sci USA, 2013.
  44. 44. 6/9/16 44 Amygdala PFC Hippocampus 0 50 100 150 200 SRprotein(%naive) Naive Sham ArcProtein(%Naive) Amygdala PFC Hippocampus 0 50 100 150 200 Naive Fear* * * SRprotein(%naive) Amygdala PFC Hippocampus 0 50 100 150 Naive Fear* 1 2 3 4 5 0 20 40 60 80 100 Interval %Freezing No Shock (Tone) Shock (Tone) No Shock (Trace) Shock (Trace) Amygdala PFC Hippocampus 0 50 100 150 200 Arcprotein(%naive) Naive Sham * A C B D E F 60#min# Day#1# 20s#tone# 2s#shock# Context#A# 20s#trace# HC# Sac# 3#min# SR and Arc are induced during consolidation of trace fear conditioning Balu et al., in preparation
  45. 45. 6/9/16 45 Amygdala PFC Hippocampus 0 50 100 150 SRprotein(%naive) Naive Sham ArcProtein(%Naive) Amygdala PFC Hippocampus 0 50 100 150 200 Fear Naive* * SRprotein(%naive) Amygdala PFC Hippocampus 0 50 100 150 Fear Naive * No Shock Shock 0 20 40 60 80 100 %Freezing Shock No Shock * A C D E F Amygdala PFC Hippocampus 0 50 100 150 200 Arcprotein(%naive) Naive Sham Day$1$ Day$2$ 20s$tone$ 2s$shock$ Context$A$ 20s$trace$ 30$min$ Sac$ HC$ 8$min$ 3$min$ B Balu et al., in preparation SR and Arc are induced after context extinction
  46. 46. 6/9/16 46 D-Cycloserine enhances extinction of fear- potentiated startle in rats through the NMDA receptor glycine modulatory site Walker et al., J Neurosci 22: 2343-2351, 2002
  47. 47. 6/9/16 47 Cognitive behavioral therapy desensitization in acrophobia significantly improved by treatment with D- cycloserine in virtual Hyatt Regency glass elevator Ressler et al., Arch Gen Psychiatry 61: 1136-1144, 2004
  48. 48. 6/9/16 48 Neurodegenerative Disorders
  49. 49. 6/9/16 49 NMDA receptor implicated broadly in neuronal degeneration by animal models 1. Stroke 2. Traumatic brain injury 3. Hypoglycemia 4. Cyanide poisoning 5. Alzheimer’s disease 6. Amyotrophic lateral sclerosis 7. Huntington’s disease 8. Epilepsy 9. Neuropathic pain Q Zhou & M Shen, Neuropharmacology 74: 68-75, 2013 Kalia et al., Lancet Neurol 2008: 742-55.
  50. 50. 6/9/16 50 SR-/- protects against middle cerebral artery occlusion infarction Mustafa et al., J Neurosci. 30: 1413, 2010
  51. 51. 6/9/16 51 Neurotoxicity is markedly reduced in SR-/- cerebral cortical cultures A. NO measured by DAF-diacetate B. nNOS and SR expressio C. Propidium staining after oxugen-glucose deprivation D. Neuronal death in WT vs SR-/-
  52. 52. 6/9/16 52 Abe et al., J Cereb Blood Flow& Metab 34: 1928-35, 2014 Confirmation of the neuroprotective effect of SR-/- in stroke model
  53. 53. 6/9/16 53 Amyotrophic Lateral Sclerosis 1. Most common adult-onset motor neuron disease 2. Life-time risk: 1 per 2000 3. Death of upper and lower motor neurons 4. 90% sporadic 5. 10% heritable 6. 2% mutations of SOD1 Rothstein et al., Ann Neurol 65 (suppl) S3-S9, 2009
  54. 54. 6/9/16 54 Paul et al., Neurobiology of Disease 35: 876-885, 2014 D-Amino acid oxidase mutation (R199W) causes familial amyotrophic lateral sclerosis (FALS) through protein misfolding and D-serine
  55. 55. 6/9/16 55 Huntington’s Disease 1. Autosomal dominant 2. Choreo-athetiod movement disorder 3. Psychiatric symptoms 4. Tri-nucleotide repeat expansion (CAG) leading to poly Q 5. Anticipation 6. Increased vulnerability of striatal GABAergic neurons to NMDARs Raymond et al., Neuroscience 198: 252-73, 2011
  56. 56. 6/9/16 56 UPF648 Elevating kynurenic acid, an endogenous NMDA receptor glycine modulatory site inhibitor, prevents neurodegeneration in a Drosophila model for Huntington’s dosease (Htt93Q) Campesan et al., Current Biology 21: 961-966, 2011
  57. 57. 6/9/16 57 Conclusion D-serine is the primary glycine site modulator on NR1 for forebrain NMDA receptors in normal function and in pathologic states.
  58. 58. 6/9/16 58 Acknowledgement Collaborators: Robbie Greene Debbye Yurgelun-Todd Don Goff John Lisman GuoChuan Tsai Richard Bergeron Toru Nishikawa Vadim Bolshakov P. Jeffery Conn Yan Li Glen Konopaske Kim Do Michel Cuenod Post-Doctoral Fellows: Alo Basu Michael Benneyworth Kazunori Suzuki Darrick Balu Shunsuke Takagi Mathew Puhl Grant Support: NIMH Conte Center (PI) NIMH RO1 (PI) NIDA R21 (PI) NARSAD
  59. 59. 6/9/16 59
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  63. 63. 6/9/16 636/9/16 63 Balu et al., Cell Mol Neurobiol 34: 419, 2014 SR PV BA4 * * Merge SR+ PV neurons in human cortex D-­Serine-­ serine SOM MergePV CA1 Serine Racemase and D-Serine are Expressed in Parvalbumin- and Somatostatin-Positive Fast- firing Cortical GABAergic Neurons
  64. 64. 6/9/16 64 Features shared between schizophrenia and bipolar disorder 1. Psychosis Goes FS et al. Curr Psychiatry Rep. 2008 Apr;10(2):178-89. 1. Ketamine as a mood elevator Krystal JH. Arch Gen Psychiatry. 2010 Nov;67(11):1110-1. 3. Cognitive impairments Burdick KE et al., Psychol Med. 2014 Oct;44(14):3083-96. 4. Cortico-limbic structural deficits Anderson D et al. Bipolar Disord. 2013 Sep;15(6):680-93. 5. Genetic risk factors: CACNA 1C Nurnberger JI Jr et al., JAMA Psychiatry. 2014 Jun;71(6):657-64.
  65. 65. 6/9/16 65 Bipolar disorder shares dendritic and spine pathology with schizophrenia Konopaske et al., JAMA Psychiatry 2014, published online, October 1.
  66. 66. 6/9/16 66 Konopaske et al., JAMA Psychiatry 71: 1323-31, 2014. Golgi-stained pyramidal neurons in layer 3 prefrontal cortex
  67. 67. Co-localztion of D-Serine in PV+ and Som+ neurons in cortev Balu et al., Cell Mol Neurobiol 34: 419, 2014
  68. 68. 6/9/16 686/9/16 686/9/16 68 0.0 0.1 0.2 0.3 * D-­Serine  (µM) Control nSR-­/-­ * 0 1 2 3 4 * Taurine(µM) Control * nSR-­/-­ 0.0 0.5 1.0 1.5 2.0 2.5 Glycine  (µM) N.S. Control nSR-­/-­ Extracellular D-serine is reduced in the hippocampus of neuronal SRKO mice • Neuronal SR-/-: 11% Reduced HP tissue D-serine content • Astrocyte SR-/-: No changes in D-serine content • Constitutive SR-/-: 90% reduced D- serine -80 -60 -40 -20 0 20 40 60 80 100120140160180 0 2 4 6 Time (min) Taurineconcentration(µM) NMDA 75µM Time  (min) NMDA  75µM Control nSRCKO Taurine(µM) Ishiwata S, Umino A, Balu DT, Coyle JT and Ishikawa T. J Neural Transm. 2015 Aug;122(8):1099-103..
  69. 69. 6/9/16 69 N-Acetl-aspartyl glutamate shows increasing inhibition of NMDA receptors with decreasing pH Klacho et al., Neurobiology of Disease 82: 580-592, 2015
  70. 70. 6/9/16 70 DA Glu Glu GABA GlyT1 inhibitor D-Serine DAAO inhibitor mGluR5 agonist TrkB agonist Ventral Striatum mGluR3 Agonists Positive GABAA modulator (alpha2) Neuroplasticity + + VTA NBM Alpha 7 nAChR Positive Modulator +

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